Review: Interpreting Antimicrobial Susceptibility Results

Susceptibility and Resistance

These are related terms to describe the same concept: the
concentration of antimicrobial required to inhibit growth of an
isolate of bacteria as it relates to the likelihood of clinical
success. "Susceptible" and "resistant" are qualitative terms
used to simplify the results of tests of growth inhibition.
Problems with interpretation can occur when the definitions
underlying the qualitative terms are not universally agreed upon or
when different thresholds are used by the individual that performs
the test and the individual that uses the results of the test.

Differentiating inherent and acquired resistance is important
from a clinical and epidemiological standpoint, since the concern
for resistance associated with treatment failure is generally
speaking due to bacteria acquiring resistance genes from other
bacteria, rather than a gene that has always been present in a
particular bacterial species. Acquired resistance is most
often generated by the acquisition of new bacterial DNA by various
mechanisms, including transfer of plasmids (extra-chromosomal DNA)
between a resistant and a susceptible organism, or transfer of an
integron or other type of moveable gene between a resistant and a
susceptible organism. Transfer of resistance can occur
between bacteria of the same species, but also between bacteria of
different species and even of different genera. The
significance of this transfer is that the bacteria containing these
resistance genes may be selectively targeted for survival by the
presence of an antimicrobial, and the resistance DNA is then
carried on the next generation or is transferred to other
bacteria.

Determining susceptibility or resistance

Since we have described resistance as requiring more
antimicrobial than can be delivered to the infection site, we need
a way to determine how much of a particular antimicrobial is
required to inhibit the growth of a particular organism. We
are all familiar with susceptibility testing as performed by most
clinical microbiology labs, but it is useful to briefly review
these tests.

The two major types of susceptibility testing performed by
veterinary diagnostic laboratories are disk diffusion and broth
microdilution. Disk diffusion testing uses paper disks
containing known quantities of antimicrobials, and the zone around
which no growth of bacteria occurs correlates with a particular
range of antimicrobial concentrations. The correlation
between zone and MIC is a qualitative rather than a quantitative
one: zones of inhibition do not linearly correspond to minimum
inhibitory concentrations of antimicrobial.

Broth microdilution testing is used to characterize the quantity
of antimicrobial required to inhibit bacterial growth.
Varying concentrations of antimicrobial are mixed with the broth
used to grow bacterial isolates, and the lowest concentration which
demonstrates no growth is the MIC. This type of testing is
usual performed with 96-well plates so multiple drugs can be tested
or even more than one isolate can be tested on a plate.
Concentrations of antimicrobial which are clustered around the
breakpoint are generally selected for testing, with the idea that
these concentrations are also actually clinical achievable in the
animal.

What is a breakpoint and how are they determined?

The purpose of the breakpoint is to provide a cutoff or
threshold for categorizing organisms with different
phenotypes. The phenotypes practicing veterinarians are
interested in identifying are those that might be difficult to
eliminate in clinical infections because they require increased
concentrations of antimicrobials to inhibit growth. However,
other groups may be interested in other cutoffs, such as
epidemiologic cutoffs, whereby populations of organisms could be
evaluated over time to watch for significant changes in
susceptibility. Different cutoffs might also be used to
evaluate whether the majority of isolates remain wild-type, or
whether acquired resistance is penetrating a population. The
breakpoints discussed in this document are clinical breakpoints,
related to predicting clinical outcome of antimicrobial therapy for
bacterial disease.

Through a multi-step process, data are gathered and/or generated
which allow the Clinical Laboratory Standards Institute Veterinary
Antimicrobial Susceptibility Testing Subcommittee (CLSI VAST) to
select appropriate breakpoints. (The CLSI is an
international, interdisciplinary, nonprofit, standards-developing,
and educational organization that promotes the development and use
of voluntary consensus standards and guidelines within the health
care community.) These data include pharmacokinetics of the drug in
question, pharmacodynamics of the drug (how does it work best), and
MIC data for at least 100 different isolates of the bacterial
species for which the breakpoints will be valid.

For a description of how tilmicosin breakpoints for porcine
respiratory disease were determined, click here, and
ceftiofur breakpoints for porcine respiratory disease, click here. A
complete review of how breakpoints for human antimicrobials are
determined can be found
here.